Abstract This I/START application, in response to PAR-15-326, will allow the PI to design and collect ?proof of concept? brain imaging data on the neural signature of prescription opiate (PO) dependence. Opiate overdose, specifically the PO overdose has become an epidemic in the U.S. This grant will help the PI develop expertise in imaging methodologies that she has not used [novel resting state functional connectivity (RSFC) analysis (modulatory interactions); the latest multiband MR acquisition protocol; diffusion tensor imaging (DTI)] to conduct a multi-method assessment of brain structural and functional alterations in prescription opiate dependent (POD) individuals vs. controls. This grant, along with PI?s expertise in her current NIDA K01 cocaine grant, will provide strong credentials for the PI and pilot data to successfully compete for her first R01 grant and conduct neuroimaging research in PO and other addiction populations by using multimodal imaging techniques. The structural and functional alterations in POD individuals vs. controls will be examined in the drug cue processing network (DCPN) consisting of structures belonging to the mesocorticolimbic and nigrostriatal systems using functional magnetic resonance imaging (fMRI) and structural, functional and effective connectivity (SC, FC, EC) analysis tools. We target the DCPN because it allows us to examine functional and structural interplay between critical brain areas in drug users that give rise to craving and compulsive drug seeking. We also will examine other large scale brain networks that overlap with DCPN. The PI and colleagues will examine group differences in brain volume and white matter connectivity using voxel based morphometry (VBM) and DTI techniques, respectively, and FC (using a resting state task) and EC (using a PO cue exposure task) among the regions within the DCPN and other large scale networks between PODs and controls (Aim 1). Sex differences in structure and function within the DCPN and other large scale networks between the groups also will be examined (Aim 2). Thirty PODs and 30 age- and gender-matched controls will be recruited. Each participant will take part in one session including two structural scans, and three functional scans (two resting state scans, and one scan during PO cue exposure task where we also will collect craving ratings). In the analysis, we will test the DCPN model for PODs, and also will examine other brain networks. Fractional Anisotropy (FA, SC measure), FC and EC strengths obtained from a series of analysis from FSL?s tract-based spatial statistics (FA values), FC values, higher order FC values (for novel RSFC analysis), and IMaGES (EC) will be analyzed using repeated measures ANCOVA utilizing group (POD, control), and sex (M, F) as between subjects factors and network (DCPN vs. other networks) as within subject measures. Similar analysis will be done using brain volume as a repeated factor. For each network, correlation between craving ratings and connectivity value in PODs will be calculated. We may identify future sex-specific targets for therapies and the identified network may be used as outcome metrics in clinical trials.